Microtubule regulation of actomyosin dynamics and force generation in T lymphocytes

T 淋巴细胞中肌动球蛋白动力学和力产生的微管调节

基本信息

批准号：
10359737
负责人：
Arpita Upadhyaya
金额：
$ 30.78万
依托单位：
UNIV OF MARYLAND, COLLEGE PARK
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-03-15 至 2023-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10359737
关键词：
APC gene Actins Actomyosin Adaptor Signaling Protein Adhesions Adhesives Antigens Binding Biochemical Biological Biomechanics CD8B1 gene Cell Communication Cell physiology Cells Characteristics Clonal Expansion Coupled Coupling Cues Cytoskeleton Cytotoxic T-Lymphocytes Development Environment Family Filament Functional disorder Future Gel Generations Goals Guanine Guanine Nucleotide Exchange Factors Human Immune response Immune system Immunologic Deficiency Syndromes Immunotherapy In Vitro Infection Inflammatory Interleukin-12 Interleukin-2 Intervention Lead Link Lymphocyte Activation Lymphoma Maintenance Mechanics Mediating Microtubules Molecular Movement Mus Mutation Myosin ATPase Optics Pathway interactions Play Plus End of the Microtubule Process Proteins Receptor Signaling Regulation Rho-associated kinase Role Signal Pathway Signal Transduction Site Small Interfering RNA Structure Surface System T-Cell Activation T-Cell Receptor T-Lymphocyte Techniques Testing Therapeutic Traction Force Microscopy Translating Work adaptive immune response base cancer cell cell killing cytokine cytotoxic CD8 T cells genetic regulatory protein human disease immunological synapse in vivo knock-down mechanical force mechanical properties mechanical stimulus mechanotransduction mutant neoplastic cell novel novel strategies optogenetics physical property polymerization quantitative imaging response sensor spatiotemporal transmission process tumor

项目摘要

Cell-cell interactions, mediated by adhesion and signaling receptors, are highly dynamic and subject to cytoskeletal movements that impart substantial mechanical force at the interface. How cells combine mechanical and biochemical signals to carry out specific functions is not well understood. Cells of the immune system present a compelling context for studying force transmission and mechanosensing because they are structurally dynamic and are sites of biochemical information transfer. T cell signaling is closely linked to the cytoskeleton, and it is evident that forces applied by the actin cytoskeleton at the T cell receptor are transduced to biochemical signaling leading to T cell activation. However, the molecular mechanisms by which these forces are regulated and how they contribute to T cell function remain obscure. Here, we propose to dissect the interactions and activities of proteins that reside at the intersection of actin and microtubule (MT) dynamics to advance our understanding of force generation and mechanosensing in T cells. We hypothesize that dynamic microtubules modulate the T cell cytoskeleton and proximal signaling both by 1) regulating actin polymerization dynamics in the lamellipodium and the assembly of structures in the lamella and 2) regulating RhoA activation leading to myosin contractility and force generation. Ultimately, we hypothesize that MT/actin interactions contribute to the ability of T cells to adapt their activation and effector function in response to the stiffness of target cells. Our first goal will be to examine the mechanisms by which MT regulate actin dynamics by probing the specific interactions between MT and actin via +TIP proteins. We will combine optogenetic techniques with mutations to probe specific interactions between MT and actin that regulate T cell activation. Our second goal will be to dissect the mechanisms that link dynamic MTs to myosin driven contractile force generation. We will combine optogenetic control of RhoA activation and inhibition with quantitative imaging and traction force microscopy to elucidate the spatiotemporal characteristics of RhoA activation during T cell activation. We will use novel sensors for GEF-H1 activity and mutations to establish its role in MT/actin coupling, force generation and T cell signaling. Finally, we will perform studies with mouse cells in a functional context to test the hypothesis that regulation of actomyosin dynamics and contractility tunes the mechanical coordination of cytotoxic T lymphocyte activation and their efficacy in killing cancer cells. Our proposed studies will clarify how mechanical stimuli and biochemical signaling are coupled during the immune response. Furthermore, the specific pathways studied in this proposal are linked to a number of immunodeficiencies and lymphoma progression and thus will help lead to a better understanding of how their dysfunction can contribute to human disease, thus providing new targets for intervention in immune therapy.

通过粘附和信号受体介导的细胞细胞相互作用是高度动态的，受在界面处赋予大量机械力的细胞骨架运动。细胞如何结合机械并且尚不清楚执行特定功能的生化信号。存在免疫系统的细胞用于研究力传输和机械传感的令人信服的环境，因为它们是结构动态的并且是生化信息转移的地点。 T细胞信号与细胞骨架紧密相关，它是显然，肌动蛋白细胞骨架在T细胞受体上施加的力被转导为生化信号传导导致T细胞激活。但是，这些力受调节的分子机制以及如何它们有助于T细胞功能保持晦涩。在这里，我们建议剖析驻留在肌动蛋白和微管（MT）动力学相交的蛋白质，以提高我们对 T细胞中的力产生和机械感应。我们假设动态微管调节T细胞细胞骨架和近端信号传导均通过1）调节lamellipodium中的肌动蛋白聚合动力学以及片段中的结构组装和2）调节RhoA激活导致肌球蛋白收缩力和力产生。最终，我们假设MT/肌动蛋白相互作用有助于T细胞的能力适应其激活和效应子功能，以响应靶细胞的刚度。我们的第一个目标是检查MT通过探测MT之间的特定相互作用来调节肌动蛋白动力学的机制和肌动蛋白通过 +尖端蛋白。我们将将光遗传技术与突变结合起来，以探测特定的相互作用在调节T细胞激活的MT和肌动蛋白之间。我们的第二个目标是剖析链接的机制动态MTS肌球蛋白驱动的收缩力产生。我们将结合RhoA的光遗传学控制用定量成像和牵引力显微镜激活和抑制，以阐明时空 T细胞激活过程中RHOA激活的特征。我们将使用新颖的传感器进行GEF-H1活动和突变以确定其在MT/肌动蛋白偶联，力产生和T细胞信号传导中的作用。最后，我们将表演在功能上下文中对小鼠细胞的研究，以检验肌动球蛋白动力学调节的假设收缩性调整了细胞毒性T淋巴细胞激活的机械协调及其在杀死癌细胞。我们提出的研究将阐明机械刺激和生化信号传导如何在免疫反应期间结合。此外，该提案中研究的具体途径与许多免疫缺陷和淋巴瘤进展，因此将有助于更好地理解它们的功能障碍如何导致人类疾病，从而为免疫干预提供新的靶标治疗。

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Cytotoxic T Lymphocyte Activation Signals Modulate Cytoskeletal Dynamics and Mechanical Force Generation.

DOI：
10.3389/fimmu.2022.779888
发表时间：
2022
期刊：
Frontiers in immunology
影响因子：
7.3
作者：
Pathni A;Özçelikkale A;Rey-Suarez I;Li L;Davis S;Rogers N;Xiao Z;Upadhyaya A
通讯作者：
Upadhyaya A

Phase separation in transcription factor dynamics and chromatin organization.

DOI：
10.1016/j.sbi.2021.06.009
发表时间：
2021-12
期刊：
Current opinion in structural biology
影响因子：
6.8
作者：
Wagh K;Garcia DA;Upadhyaya A
通讯作者：
Upadhyaya A

Nanotopography modulates cytoskeletal organization and dynamics during T cell activation.

DOI：
10.1091/mbc.e21-12-0601
发表时间：
2022-09-01
期刊：
MOLECULAR BIOLOGY OF THE CELL
影响因子：
3.3
作者：
Wheatley, Brittany A.;Rey-Suarez, Ivan;Hourwitz, Matt J.;Kerr, Sarah;Shroff, Hari;Fourkas, John T.;Upadhyaya, Arpita
通讯作者：
Upadhyaya, Arpita

Three-dimensional structured illumination microscopy with enhanced axial resolution.

DOI：
10.1038/s41587-022-01651-1
发表时间：
2023-09
期刊：
NATURE BIOTECHNOLOGY
影响因子：
46.9
作者：
Li, Xuesong;Wu, Yicong;Su, Yijun;Rey-Suarez, Ivan;Matthaeus, Claudia;Updegrove, Taylor B. B.;Wei, Zhuang;Zhang, Lixia;Sasaki, Hideki;Li, Yue;Guo, Min;Giannini, John P. P.;Vishwasrao, Harshad D. D.;Chen, Jiji;Lee, Shih-Jong J.;Shao, Lin;Liu, Huafeng;Ramamurthi, Kumaran S. S.;Taraska, Justin W. W.;Upadhyaya, Arpita;La Riviere, Patrick;Shroff, Hari
通讯作者：
Shroff, Hari